Amyloid-B peptide (AB) is believed to have a central role in the pathogenesis of Alzheimer's disease (AD). Intracellular mechanisms by which AB disrupts cellular properties, leading to neuronal and synaptic dysfunction, and mitochondrial damage, are not well understood. ABAD (A6 Binding Alcohol Dehydrogenase) is an intracellular enzyme with unique features including a wide range of substrates, binding AB in the nanomolar range, and exacerbating AB-induced cell stress. Recent results from this laboratory and others support a connection between ABAD and AB-mediated mitochondrial and synaptic dysfunction (project 3) relevant to AD pathology. Ap interacts specifically with ABAD in the mitochondria of AD patients and transgenic mice. Transgenic mice overexpressing ABAD in an Ap-rich milieu present exaggerated neuronal oxidative stress, mitochondrial dysfunction, and impaired memory. The crystal structure of Ap-bound ABAD shows substantial deformation of the active site that prevents nicotinamide adenine dinucleotide (NAD) binding. From the crystal structure, the putative AB interaction site in ABAD spans residues 94-114. A peptide encompassing this region of ABAD, residues 91-119, specifically inhibits the ABAD-Ap interaction and suppresses Ap-induced apoptosis and free- radical generation in neurons. Indicating that binding of ABAD to AB is crucial in AB-mediated neuronal stress. Possible mechanisms underlying ABAD-mediated changes in mitochondrial properties are indicated by our findings that AB inhibits ABAD enzymatic activity, the observation that ABAD binds cyclophilin (cypD), sequestering the latter in the mitochondrial matrix, and potential toxic effects of ABAD-ABcomplex (e.g., generation of ROS). This competitive renewal is founded on the hypothesis that ABAD is a critical intracellular target potentiating AB-mediated mitochondrial dysfunction and, thus, leading to cellular perturbation in an AB-rich environment. Blockade of the ABAD-ABinteraction may be a potential therapeutic approach in the treatment of AD. The aims of Project 4 are to identify the basis of ABAD-AB, ABAD-cyclophilin D (cypD) interactions using crystal structure techniques, and to analyze critical aspects of the intracellular pathway through which ABAD engages AB, cypD, and ABAD-AB complex induces cellular stress. Project 4 will work closely with Projects 1-3, and will obtain technical assistancefrom CoreA. Collaborative interactions will include: sharing of reagents for analysis of ABAD (Project 3), mechanisms for evaluating cell stress (Projects 1-3), and assistance in data analysis (core A).